Thermodynamic study of yeast phosphoglycerate kinase.


Abstract

Enthalpies of binding of MgADP, MgATP, and 3-phosphoglycerate to yeast phosphoglycerate kinase have been determined by flow calorimetry at 9.95-32.00 degrees C. Combination of these data with published dissociation constants [Scopes, R.K. (1978) Eur. J. Biochem. 91, 119-129] yielded the following thermodynamic parameters for the binding of 3-phosphoglycerate at 25 degrees C: delta Go = -6.76 +/- 0.11 kcal mol-1, delta H = 3.74 +/- 0.08 kcal mol-1, delta So = 35.2 +/- 0.6 cal K-1 mol-1, and delta Cp = 0.12 +/- 0.32 kcal K-1 mol-1. The thermal unfolding of phosphoglycerate kinase in the absence and presence of the ligands listed above was studied by differential scanning calorimetry. The temperature of half-completion, t 1/2, of the denaturation and the denaturational enthalpy are increased by the binding of the ligands, the increase in t 1/2 being a manifestation of Le Chatelier's principle and that in enthalpy reflecting the enthalpy of dissociation of the ligand. Only one denaturational peak was observed under all conditions, and in contrast with the case of yeast hexokinase [Takahashi, K., Casey, J.L., & Sturtevant, J.M. (1981) Biochemistry 20, 4693-4697], no definitive evidence for the unfolding of more than one domain was obtained. Study holds ProTherm entries: 3909, 3910, 3911, 3912, 3913, 3914, 3915, 3916 Extra Details: dissociation constants; binding; ligands; domain

Submission Details

ID: KWg5e7EC

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:23 p.m.

Version: 1

Publication Details
Hu CQ;Sturtevant JM,Biochemistry (1987) Thermodynamic study of yeast phosphoglycerate kinase. PMID:3548815
Additional Information

Number of data points 32
Proteins Phosphoglycerate kinase ; Phosphoglycerate kinase
Unique complexes 1
Assays/Quantities/Protocols Experimental Assay: dCp pH:9.0, buffers:borate: 50 mM ; Experimental Assay: dHcal pH:9.0, buffers:borate: 50 mM ; Experimental Assay: Tm pH:9.0, buffers:borate: 50 mM ; Experimental Assay: dHvH pH:9.0, buffers:borate: 50 mM ; Experimental Assay: dCp pH:8.5, buffers:borate: 50 mM ; Experimental Assay: dHcal pH:8.5, buffers:borate: 50 mM ; Experimental Assay: Tm pH:8.5, buffers:borate: 50 mM ; Experimental Assay: dHvH pH:8.5, buffers:borate: 50 mM ; Experimental Assay: dCp buffers:PIPES: 50 mM, pH:7.96 ; Experimental Assay: dHcal buffers:PIPES: 50 mM, pH:7.96 ; Experimental Assay: Tm buffers:PIPES: 50 mM, pH:7.96 ; Experimental Assay: dHvH buffers:PIPES: 50 mM, pH:7.96 ; Experimental Assay: dCp pH:7.46, buffers:PIPES: 50 mM ; Experimental Assay: dHcal pH:7.46, buffers:PIPES: 50 mM ; Experimental Assay: Tm pH:7.46, buffers:PIPES: 50 mM ; Experimental Assay: dHvH pH:7.46, buffers:PIPES: 50 mM ; Experimental Assay: dCp pH:7.04, buffers:PIPES: 50 mM ; Experimental Assay: dHcal pH:7.04, buffers:PIPES: 50 mM ; Experimental Assay: Tm pH:7.04, buffers:PIPES: 50 mM ; Experimental Assay: dHvH pH:7.04, buffers:PIPES: 50 mM ; Experimental Assay: dCp buffers:PIPES: 50 mM, pH:6.5 ; Experimental Assay: dHcal buffers:PIPES: 50 mM, pH:6.5 ; Experimental Assay: Tm buffers:PIPES: 50 mM, pH:6.5 ; Experimental Assay: dHvH buffers:PIPES: 50 mM, pH:6.5 ; Experimental Assay: dCp pH:5.88, buffers:PIPES: 50 mM ; Experimental Assay: dHcal pH:5.88, buffers:PIPES: 50 mM ; Experimental Assay: Tm pH:5.88, buffers:PIPES: 50 mM ; Experimental Assay: dHvH pH:5.88, buffers:PIPES: 50 mM ; Experimental Assay: dCp pH:5.4, buffers:acetate: 50 mM ; Experimental Assay: dHcal pH:5.4, buffers:acetate: 50 mM ; Experimental Assay: Tm pH:5.4, buffers:acetate: 50 mM ; Experimental Assay: dHvH pH:5.4, buffers:acetate: 50 mM
Libraries Mutations for sequence SLSSKLSVQDLDLKDKRVFIRVDFNVPLDGKKITSNQRIVAALPTIKYVLEHHPRYVVLASHLGRPNGERNEKYSLAPVAKELQSLLGKDVTFLNDCVGPEVEAAVKASAPGSVILLENLRYHIEEEGSRKVDGQKVKASKEDVQKFRHELSSLADVYINDAFGTAHRAHSSMVGFDLPQRAAGFLLEKELKYFGKALENPTRPFLAILGGAKVADKIQLIDNLLDKVDSIIIGGGMAFTFKKVLENTEIGDSIFDKAVGPEIAKLMEKAKAKGVEVVLPVDFIIADAFSASANTKTVTDKEGIPAGWQGLDNGPESRKLFAATVAKATVILWNGPPGVFEFEKFAAGTKALLDEVVKSSAAGNTVIIGGGDTATVAKKYGVTDKISHVSTGGGASLELLEGKELPGVAFLSEKK
Sequence Assay Result Units